Effect of unripe banana flour as a functional feed ingredient on growth performance, internal organ relative weight and carcass traits of broilers

Abstract Background Following the prohibition of in‐feed antibiotics, poultry nutritionists are increasingly interested in the use of functional feed. Unripe banana flour (UBF) contains significant amounts of oligosaccharides (which may act as prebiotics) and antioxidants, making it a potential functional feed for broilers. However, research on the use of UBF as a functional feed ingredient for broilers is limited. Objectives The study investigated the effect of UBF with or without probiotic and multienzyme on growth, internal organ weight and carcass characteristics of broilers. Methods A total of 392 broiler chicks were distributed into 4 groups included CONT (chicks receiving control feed), UBF (chicks receiving 5% UBF in feed), UBFPRO (5% UBF plus 0.05% probiotics) and UBFZYM (5% UBF plus 0.05% multienzyme). Data on growth performance were weekly recorded, whereas data on internal organs and carcass were collected on day 38. Results Feed conversion ratio (FCR) was lower (p < 0.05) in UBF, UBFPRO and UBFZYM than that in CONT chicks, with no significant difference in body weight, body weight gain and feed intake. There was a tendency that gizzard was higher (p = 0.08) in CONT than in UBF chicks. Also, pancreas tended (p = 0.09) to be lower in UBFZYM than in CONT birds. There was a notable effect (p < 0.05) of dietary treatments on the yellowness (b*) values of thigh meats, in which UBFPRO had lower b* values than that of CONT but did not differ from that of UBF and UBFZYM. There was no difference (p < 0.05) in carcass and commercial proportion of broilers. Conclusions Feeding of 5% UBF with or without probiotic and multienzyme improved FCR, without negatively affecting the carcass characteristics of broilers.


INTRODUCTION
Broiler chicken is one of the most important livestock commodities for supplying animal protein to people. Broiler industry is also crucial to the economy of many countries. Hence, the sustainability of the broiler farming is very critical for the fulfilment of nutrition as well as the economy of the country. Antibiotic growth promoters (AGP) have widely been utilized in the broiler industry for decades to maximize growth potential and health. However, most countries, including Indonesia, have outlawed the use of AGP for food safety reasons. Indeed, excessive and continuous use of AGP in broiler chickens can result in AGP residues in the meat, posing a health risk to consumers (Sugiharto, 2016). The use of functional feed is increasingly attracting the attention of poultry nutritionists following the prohibition of AGP on broiler production. Besides containing nutrients as conventional feedstuffs, functional feed contains various bioactive compounds that can have promoting impacts on the health and growth of broilers (Sugiharto et al., 2018). Moreover, functional feed could improve the carcass traits (Alwaleed et al., 2020) and meat quality of broilers (Nopparatmaitree et al., 2022).
Study has shown that unripe (green) banana flour, in addition to having a high energy (Dumorné et al., 2020), contains substantial amount of oligosaccharides and resistant starch (Chang et al., 2022).
These active components are often attributed to prebiotics, which can improve intestinal ecology and function of poultry. Unripe banana flour (UBF) also showed a high antioxidant activity (Chang et al., 2022;Padam et al., 2014), which is essential in improving the physiological condition and health of broilers reared under intensive system. In line with the above studies, our recent study showed that four cultivars of UBF in Indonesia contained energy exceeding 3000 kcal/kg (Munita et al., 2022). These data confirm that UBF can be used as an alternative energy source to replace yellow corn, the price of which is very volatile.
Our data also showed that UBF has prebiotic activity that can support the growth of probiotic bacteria (Lactobacillus casei). Also, UBF showed a high antioxidant activity, as indicated by the ability of UBF in scavenging the 2,2-diphenyl-1-picrylhydrazyl (DPPH) (Munita et al., 2022). The use of UBF as a feed ingredient for broilers is currently still lacking. To the best of our knowledge, only Dumorné et al. (2020) used UBF as the energy source for broiler chickens. They confirmed that the inclusion of banana flour into broiler feeds up to 20% had no adverse effect on broiler growth performance.
UBF is generally produced from the discarded/culled green bananas during fruit selection and processing. In some banana-producing countries, culled green bananas are so abundant that, if not utilized, they can cause environmental problems (Padam et al., 2014). In addition to banana cultivars commonly consumed by humans, several types of bananas are not favoured by consumers because they have a sour and unpleasant taste. As a consequence, these cultivars of banana have low economic value (Munita et al., 2022). Taking these above facts into consideration, the application of UBF as functional feed ingredients may therefore increase the economic worth of the underconsumed banana cultivars as well as support the sustainability of broiler production. Probiotics and enzymes are two types of feed additives that have widely been applied to broiler chicken production. Besides being able to work individually, probiotics are reported to be able to synergize with prebiotics to further improve intestinal function and growth performance of broiler chickens (Sugiharto, 2016). The synergistic effect was also seen between prebiotics and enzymes in improving broiler production performance as confirmed by Shang and Kim (2017). In this study, UBF was included into broiler feeds in conjunction with probiotic L. casei and multienzyme. The aim of the present study was to investigate the effect of UBF as a functional feed ingredient with or without probiotic and multienzyme on growth performance, internal organ relative weight and carcass characteristics of broilers.

Preparation of unripe banana flour
The underconsumed banana cultivar ("Norowito" cultivar), which has a sour and unpleasant taste (Munita et al., 2022), was used to pro- was also used in this study.
The chicks were raised in a rice husk-bedded open-sided broiler house. Throughout the trial, a constant lighting schedule was applied.
The feeds (mash form) were prepared to be isocaloric and isonitrogenous, and they met the Indonesian National Standard for broiler starter (days 8-21; Table 2) and finisher feed (days 22-38; Table 3). The chicks were given Newcastle disease (ND) and infectious bronchitis vaccines by spray soon after hatching. The ND vaccination was also given to the chicks at the age of 18. Body weight (individually weighed), feed intake and feed conversion ratio (FCR) of broilers were recorded weekly. Feed intake was measured as the difference between the amount of feed

Internal organ weights of broiler chickens
There was no substantial difference (p < 0.05) among the groups of chicks with regard to the relative weight of internal organs of broilers (Table 6). Yet, there was a tendency that gizzard was higher (p = 0.08) in CONT than in UBF chicks. Also, pancreas tended (p = 0.09) to be lower in UBFZYM than in CONT birds. Contrast analysis (Table 7) showed that gizzard relative weight was different among CONT vs.

Carcass traits, pH values and colour of broiler meats
The data on carcass and commercial cuts of broilers are presented in Table 8. There was no difference (p > 0.05) between carcass and commercial proportion of broilers. However, contrast analysis (Table 9) showed the difference (p < 0.05) in eviscerated carcass between UBF and UBFPRO groups.
There was a notable effect (p < 0.05) of dietary treatments on b* values of thigh meats, in which UBFPRO had lower b* values than that of CONT but did not differ from that of UBF and UBFZYM (  , 2021). These functional effects of enzymes may therefore support the prebiotic effect of UBF in improving the nutrient utilization of broilers. In line with this present study, Shang and Kim (2017) also confirmed that prebiotics can work synergistically with enzymes in improving broiler production performance. In contrast to enzymes, the application of probiotics in combination with UBF did not exert any additional effect on the growth of broilers when compared with the application of UBF alone. So far, the definite reason for such a condition remains unclear. However, the inconsistent effect of probiotics (Soumeh et al., 2021;Sugiharto, 2016) seemed to explain the lack of synergistic effect of probiotics and UBF on broiler performances in this current study.
Data in our present study showed a clear tendency that feeding UBF reduced the relative weight of gizzard of broilers. Previous study suggested that the high content of insoluble fibre in diet may implicate further grinding activity leading to further muscular layer development and hence increasing gizzard size (Rungcharoen et al., 2013).
Taken the latter study into consideration, the high content of soluble fibre (Falcomer et al., 2019) and carbohydrates (Munita et al., 2022) in UBF may therefore reduce grinding activity and thus prevent the further muscular layer development of gizzard. Different from the UBF and UBF plus probiotics, the use of multienzyme in combination with UBF did not exert lowering effect on gizzard weight. Indeed, contrast analysis showed that UBFZYM chicks had higher gizzard weight as compared to UBF chicks. In line with our study, Amerah et al. (2015) reported that enzyme (endoxylanase and β-glucanase) supplementation increased the relative weight of gizzard of broilers in their study.
In general, enzyme has been reported to improve the digestibility of nutrients, including fibre, and thereby reduce the grinding activity and gizzard weight (Rungcharoen et al., 2013). In this regard, the attribution of multienzyme to increasing the relative weight of gizzard seemed to be inappropriate. Yet, one study by Bedford and Schulze (1998) revealed that xylanase indirectly stimulates the function of the gizzard and proventriculus (Bedford & Schulze, 1998). Owing to this, it is tempting to speculate that exogenous enzyme supplementation may enhance the growth and development (muscular layer development) of gizzard irrespective of the grinding activity of the gizzard as discussed previously.
Our current study revealed that the relative weight of pancreas activities of pancreatic enzymes, as a consequence, less quantities of enzyme are needed for the chemical digestion of feeds. The latter condition seemed to be attributed to the reduced pancreatic tissue activity (in producing pancreatic enzymes), thus lowering pancreas organ weight. It is interesting to see in this study that the use of multienzyme in combination with UBF resulted in the most substantial reduction in pancreas weight as compared to that of control. In accordance with our data, the relative weight of pancreas decreased when the exogenous glucanase (Moran, 1985) or protease (Nastain et al., 2021) was supplemented to the diets of broilers. It was most likely that exogenous enzymes improved chemical digestion process in the gastrointestinal tract, while reducing the activity of the pancreas in producing the respective enzymes (Moran, 1985;Nastain et al., 2021).
The less activity of pancreas may thus be attributed to the reduced pancreatic tissue weight.
Data in our present study showed no substantial effect of dietary treatments on the carcass traits of broiler chickens. However, contrast analysis showed that UBFPRO had lower eviscerated carcass than UBF group, suggesting the lowering effect of probiotics on the carcass weight of broilers. Our finding was in contrast to most of the studies revealing the enhancing effect (Parsa et al., 2018) or no effect (Tang et al., 2021) of probiotics on eviscerated carcass of broilers. However, our finding was in line with Rehman et al. (2020) showing the reduced dressing percentage of broilers with the administration of commercial Abbreviations: CONT, chicks receiving control feed; UBF, chicks receiving 5% unripe banana flour in feed; UBFPRO, chicks receiving 5% unripe banana flour in feed plus 0.05% probiotics; UBFZYM, chicks receiving 5% unripe banana flour plus 0.05% multienzyme.
exogenous enzymes from the PSE condition on broiler meats. In agreement with our inference, Abdurrahman et al. (2016) reported that probiotic (Lactobacillus sp.) treatment decreased the yellowness values of broiler meats. So far, the mechanism by which the probiotics may reduce the yellowness of broilers meats remains unclear, but Abdurrahman et al. (2016) confirmed that probiotics may lower the fat content in meats, resulting in less yellow of broiler meats. In accordance with probiotics, Szymczyk et al. (2007) showed the reduced fat content in broiler meats with exogenous enzyme administration. The latter condition may therefore be responsible for the lower yellowness values of meat in UBFZYM group. With regard to the prebiotic effect of UBF on the yellowness values of thigh meat, Tavaniello et al. (2018) did not see any substantial effect of prebiotics on the lightness and yellowness indexes of broiler meats in their study.

CONCLUSIONS
Dietary inclusion of 5% UBF with or without probiotic and multienzyme improve FCR, without negatively affecting the carcass characteristics of broilers.